Female athletes injure their Anterior Cruciate Ligaments (ACLs) more frequently than male athletes in similar sports. Tissue remodeling in response to mechanical stresses determines the size and biomechanical properties of ligaments. Sex hormones affect remodeling of various tissues by regulating expression of genes encoding extracellular matrix components and the degradative enzymes that mediate matrix turnover. We hypothesize that expression of tissue remodeling genes in the ACL varies by gender. The objective of this project is to characterize differential expression of tissue remodeling genes in the human ACL. Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) control the rate of matrix degradation. Preliminary studies detected expression of nine MMPs and four TIMPs in the human ACL. These experiments also identified the first known gender difference in expression of the remodeling enzymes MMP3 (stromelysin) and MMP 1 (collagenase), and the structural protein collagen 1-alpha1 in the ACL. In the proposed studies, ACL mRNAs encoding tissue remodeling genes will be quantified. Initial targets will be the nine MMPs and four TIMPs known to be expressed in the ACL, as well as Type 1 collagen and elastin, the major structural components of the ACL. Levels of each target in male and female ACLs will be compared to detect gender differences in expression. Expression profiling using gene arrays will identify additional differentially expressed genes that may affect ACL remodeling. Effects of sex steroids on expression of remodeling genes will be assessed by culturing ACL in hormone-supplemented media that mimic the menstrual cycle. MMP and TIMP gene products will be localized within the ACL to determine their cellular origins and ability to diffuse within the ligament. Gene expression results will be corroborated by enzyme assays and Western blotting to determine activities and amounts of MMP and TIMP proteins. Results of these studies will lead to a better understanding of ACL remodeling and will improve our ability to prevent and to treat ACL injuries. Such capabilities are especially important because of the large numbers of ACL injuries that result from increasing participation of women in competitive sports.